Microchip-based capillary electrophoresis of human serum proteins.

The separation and relative quantitation of human serum proteins is important to the clinical diagnosis of various states of disease. Microchip-based capillary electrophoresis (CE) of human serum proteins offers several advantages over sodium dodecyl sulfate-poly(acrylamide) gel electrophoresis and conventional CE methods, including decreased sample consumption and analysis time and the possibility of on-chip sample manipulation (dilution, labelling, etc.). The microchip used in these studies was designed to allow for on-chip, post-separation labelling of the proteins and subsequent laser-induced fluorescence detection. 2-Toluidinonaphthalene-6-sulfonate (TNS) is a virtually non-fluorescent reagent which, upon non-covalent association with the protein and excitation at 325 nm, produces a fluorescent product with an emission maximum near 450 nm. After optimization of buffer conditions (100 mM borate with 2 mM lactate, pH 10.5), individual serum proteins (IgG to mimic the gamma zone, transferrin the beta zone, alpha-1-antitrypsin the alpha 1 zone and albumin its own zone) were successfully resolved on-chip, as was a "synthetic" serum solution composed of a mixture of all four of the previously mentioned proteins. Analysis of all five protein zones in a true human serum sample, however, has not yet been achieved on-chip due to the poor sensitivity of the TNS label for several of the serum proteins.